Regulation of KCC2 and NKCC during development: membrane insertion and differences between cell types.
The developmental switch of GABA's action from excitation to inhibition is likely due to a change in intracellular chloride concentration from high to low. Here we determined if the GABA switch correlates with the developmental expression patterns of KCC2, the chloride extruder K+-Cl- cotransporter, and NKCC, the chloride accumulator Na+-K+-Cl- cotransporter. Immunoblots of ferret retina showed that KCC2 upregulated in an exponential manner similar to synaptophysin (a synaptic marker). In contrast, NKCC, which was initially expressed at a constant level, upregulated quickly between P14 and P28, and finally downregulated to an adult level that was greater than the initial phase. At the cellular level, immunocytochemistry showed that in the inner plexiform layer KCC2's density increased gradually and its localization within ganglion cells shifted from being primarily in the cytosol (between P1-13) to being in the plasma membrane (after P21). In the outer plexiform layer, KCC2 was detected as soon as this layer started to form and increased gradually. Interestingly, however, KCC2 was initially restricted to photoreceptor terminals, while in the adult it was restricted to bipolar dendrites. Thus, the overall KCC2 expression level in ferret retina increases with age, but the time course differs between cell types. In ganglion cells the upregulation of KCC2 by itself cannot explain the relatively fast switch in GABA's action; additional events, possibly KCC2's integration into the plasma membrane and downregulation of NKCC, might also contribute. In photoreceptors the transient expression of KCC2 suggests a role for this transporter in development.